1. Field of the Invention
The present invention relates to a process for producing 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) including salts and hydrates thereof, macrocyclic compounds comprising metal ions complexes thereof and compositions comprising said macrocyclic compounds, which can be used to produce contrast agents for magnetic resonance imaging.
2. Description of the Related Art
Magnetic resonance imaging (MRI) is a powerful, non-invasive technique used to produce detailed two or three-dimensional anatomical images of tissues in the body. Conventional MRI uses the proton 1H as its signal source which is highly abundant in tissues and it has the highest sensitivity of all the biologically relevant nuclei.
Contrast, which makes the differentiation of internal structures possible in the image, arises from how the signal decays and is the difference between the resulting signals from two tissue regions. The route by which the protons release the energy they absorbed from the radio-frequency pulse, thus reducing the transverse magnetisation and causing signal decay, is known as relaxation. In MRI two independent relaxation processes occur simultaneously: spin-lattice or longitudinal relaxation characterised by the time constant T1, and spin-spin or transverse relaxation, characterised by the time constant T2.
Often, when suitable T1- or T2-weighting sequences are used, the natural contrast between two tissues is enough to produce a diagnostically-useful image. However, some conditions do not lead to specific enough changes in the relaxation times of the affected tissue though and then a contrast agent is used to locally change the relaxation times of the diseased tissue, improving the image contrast.
Most contrast agents work by shortening the relaxation times of the water protons in the targeted tissue. T1 contrast agents are based on paramagnetic metal ion chelates which make the tissue appear brighter on the T1-weighted image (positive contrast). T2 contrast agents are usually superparamagnetic iron oxide nanoparticles which create dark spots on the T2-weighted image (negative contrast). T1 agents are the most widely used and the majority of these are based on chelates of the gadolinium ion (Gd3+).
To be an effective T1 agent the gadolinium (III) chelate must significantly increase the proton relaxation rates in water. Gadolinium is the seventh element in the lanthanide series and, like the other lanthanide elements, it is most commonly found in the +3 oxidation state, corresponding to the electronic configuration [Xe]4f7. This means that Gd3+ has seven unpaired electrons, making it highly paramagnetic i.e. Gd(III) ions have large permanent magnetic moments (due to electron spin angular momentum), but in the absence of an external magnetic field these are randomly oriented. Due to its large size the Gd(III) ion typically has a coordination number of nine in its complexes. As a free ion gadolinium is very toxic for the tissues but is generally regarded as safe when administrated as a chelated compound.
The level of toxicity depends on the strength of the chelating agent, also known as ligand, chelator or sequestering agent.
Usually these ligands are organic compounds which form two or more separate coordinate bonds with a single central metal ion, in this case, the gadolinium ion, inactivating it thus reducing or eliminating its toxic effect in the tissues.
Polyaminopolycarboxylic acid compounds are the ligand type of choice because they form exceptionally stable complexes with the Gd(III) ion, which can be explained by a number of reasons. These compounds can be linear (such as pentetic acid or diethylene triamine pentaacetic acid also named as DTPA) or macrocyclic (such as 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid, DOTA). DOTA is used as the ligand in the synthesis of the MRI contrast agent gadoterate meglumine ([Gd(DOTA)(H20)](meglumine)).
Several synthetic routes for the production of DOTA have been proposed, namely by Stetter, Hermann; Wolfram Frank (1976)—“Complex Formation with Tetraazacycloalkane-N,N′,N″,N″′; —tetraacetic Acids as a Function of Ring Size”. Angewandte Chemie International Edition in English 15 (11): 686), by R. Delgado & J. J. Fraústo da Silva—Talanta, Vol. 29, pp. 815-822, Issue 10, 1982, and by J. F. Desreux—Inorg. Chem. 1980, 19, pp. 1319-1324.
The preparation of DOTA was first reported in 1976 by Stetter & Frank (full ref. above) through the reaction of 1,4,7,10-tetraazacyclododecane with chloroacetic acid in aqueous alkali medium to obtain DOTA wherein the resulting inorganic salts were separated and purified by treatment with an ion-exchange column Dowex 2×8.
The method most widely reported in the literature is typified by Delgado et al. (full ref. above), where cyclen is reacted with chloroacetic acid under aqueous basic conditions (pH =˜10) to form DOTA, which is crystallised by acidifying the cooled DOTA solution to pH 2 with hydrochloric acid and placing it in the refrigerator overnight.
Desreux (full ref. above) also reported a similar procedure, but specified sodium hydroxide as being the base used, with a reaction temperature of 80° C., and stated that upon acidification DOTA precipitates out of solution at pH 2.5.
E. Clarke & A. Martel (1991)—Inorganica Chimica Acta, 190, pp 27-36), describes the preparation of DOTA by alkylation of cyclic tetraamine ligands with bromoacetic acid at a controlled pH between 11.2 to 11.3 being the resulting product recovered by treatment with a ion-exchange column as ammonium salts followed by treatment with a potassium cation solution at pH of 11.5 and vacuum concentration. The resulting ligands were then reprotonated by addition of HCl and isolated by recrystallization from hot water.
WO9905128 discloses a process for producing DOTA compounds by 2 step-alkylation wherein the alkylation agent is preferably bromoacetic acid but also includes chloroacetic acid, in aqueous solution at a basic pH with an excess of said alkylation agent, followed by hydrolysis and purification with ion exchange resins and with an optional recrystallization step in order to obtain highly purified DOTA compounds. In particular, WO9905128 discloses a multistep process for the preparation of DOTA starting from:                a) an alkylation reaction of a 2a,4a,6a,8a-decahydrotetraazacyclopent [fg]acenaphthylene with an acid in aqueous solution and at a basic pH, followed by        b) a second alkylation reaction with a different alkylating agent, and by        c) the hydrolisis of any ester groups, and        
wherein the amount of the first alkylating agent used in step a) varies between 2-2.3 mol of reagent per mol of substrate and from 2-3 mol in step b) and the reaction temperature varies from room temperature to 80° C., depending on the reactivity of the alkylating agent.
To be able to be eventually used as a suitable contrast agent comprising gadoterate meglumine, the concentrations of process impurities present in the raw DOTA (both organic and the inorganic) must be removed or significantly reduced. This is so that the purified DOTA meets the strict specifications for use in a contrast agent or else it will not be approved for sale by the relevant medicine regulatory body as it will not be considered safe enough for human use. Therefore a series of purification steps must be employed to remove these impurities without introducing too high a concentration of a new impurity or residual solvent, as these must also meet the specifications.
However, the DOTA resulting from the above mentioned processes is still highly contaminated with organic and inorganic impurities, in particular with chloride and sodium ions, and the conventional purification steps using ion-exchange resins, as disclosed above, only solves this problem in some extent.
In fact, G. Hernandez, M. F. Tweedle and R. G. Bryant, Inorg. Chem., 1990, 29, 5109-5113, disclose the synthesis of the sodium salt of [Gd(DOTA)(H2O)]− (Na[Gd(DOTA)(H2O)].4H2O). However, this compound is unsuitable for use as a contrast agent as it contains sodium. Nevertheless, the synthetic procedure herein disclosed highlights that high temperatures (90° C.) and long reaction times (6.5 hours) are required to successfully react DOTA and gadolinium oxide (Gd2O3, an ionic salt which is the source of the gadolinium ion) together to form the thermodynamically stable [Gd(DOTA)(H2O)]−. This can be accounted for by the very slow kinetics of formation of the complex.
It is thus desirable to obtain an optimized process for the raw DOTA synthesis which ensures not only high yields of this compound, at least 50% relative to the amounts of starting reagents used, but also that the raw DOTA is of a suitable quality and in a form that was easy to work with. Furthermore, it is also desirable to simplify the method for producing DOTA at an industrial scale.